The invention discloses a preparation method of an electrochemical biosensor. The present invention first prepares a novel two-dimensional nano-electrode material FeTiO2 / MoS2, that is, a two-dimensional nano-composite material in which iron-doped titanium dioxide nano-squares are in-situ compounded with molybdenum disulfide, and utilizes the good biocompatibility and Large specific surface area, loaded with citrinin antibody, during detection, because iron-doped titanium dioxide can catalyze hydrogen peroxide to generate O2 in situ, generate electrochemical signal, and then use the specific quantitative combination of antibody and antigen to determine the electron transport ability Therefore, the current intensity is reduced accordingly, and the construction of a biosensor for detecting citrinin by a label-free electrochemical method is finally realized.